Inhibition ELISA method for detecting foot-and-mouth disease virus antibody and application

A foot-and-mouth disease virus, anti-foot-and-mouth disease virus technology, applied in the biological field, to achieve the effect of easy operation

Active Publication Date: 2021-06-01
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] In order to solve the problem of how to optimize and innovate existing methods to establish a new method for the detection of foot-and-mouth disease antibodies, the first object of the present invention is to provide a method for inhibiting the detection of foot-and-mouth disease virus antibodies. The detection kit, purpose three is to provide the application of said inhibition ELISA in the detection of foot-and-mouth disease virus antibody

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Embodiment 1 Utilizes the establishment and application of the method for measuring O-type foot-and-mouth disease antibody by inhibition ELISA

[0050] 1 Materials and methods

[0051] 1.1 Preparation of foot-and-mouth disease virus antigen and antiserum

[0052] 1.1.1 Preparation of FMDV antigen and purification of 146S antigen: Mix inactivated foot-and-mouth disease vaccine (O / Mya98 strain) and demulsifier in a certain proportion, shake and layer, centrifuge at 3000rpm, 4°C for 10 minutes, and carefully absorb the lower layer of antigen solution. Take 100ml of antigen solution, 45000rmp, ultracentrifuge at 4°C for 1h, resuspend the sediment on the bottom wall of the centrifuge tube to 2ml with PBS with a pH value of 7.6, pass through a sucrose density gradient (45%, 35%, 25%, 15%) at 35000rpm, 4 Centrifuge at ℃ for 2.5 hours, take 0.5ml samples and combine the peaks of the same order, and detect with a 260nm ultraviolet spectrophotometer, collect the absorption peak ...

Embodiment 2

[0099] Example 2 Establishment and application of the method for measuring Asian type 1 foot-and-mouth disease antibody by inhibition ELISA

[0100] 1 Materials and methods

[0101] 1.1 Preparation of foot-and-mouth disease virus antigen and antiserum

[0102] 1.1.1 Preparation of FMDV antigen and purification of 146S antigen: Mix inactivated foot-and-mouth disease vaccine (JSL2006 strain) and demulsifier in a certain proportion, shake and layer, centrifuge at 3000rpm, 4°C for 10 minutes, and carefully absorb the lower layer of antigen solution. Take 100ml of antigen solution, 45000rmp, ultracentrifuge at 4°C for 1h, resuspend the sediment on the bottom wall of the centrifuge tube to 2ml with PBS with a pH value of 7.6, and centrifuge at 35000rpm for 2.5 Hours, 0.5ml samples were taken and the peaks of the same level were combined, and detected by a 260nm ultraviolet spectrophotometer, and the absorption peak (the second peak) containing the 146S antigen of the complete virio...

Embodiment 3

[0149] Example 3 Establishment and Application of the Method for Determination of Type A Foot-and-Mouth Disease Antibody by Inhibition ELISA

[0150] 1 Materials and methods

[0151] 1.1 Preparation of foot-and-mouth disease virus antigen and antiserum

[0152] 1.1.1 Preparation of FMDV antigen and purification of 146S antigen: Mix inactivated foot-and-mouth disease vaccine (AF72 strain) and demulsifier in a certain proportion, shake and stratify, centrifuge at 3000rpm, 4°C for 10 minutes, and carefully absorb the lower layer of antigen solution. Take 100ml of antigen solution, 45000rmp, ultracentrifuge at 4°C for 1h, resuspend the sediment on the bottom wall of the centrifuge tube to 4ml with PBS with a pH value of 7.6, and centrifuge at 35000rpm for 2.5 Hours, 0.5ml samples were taken and the peaks of the same level were combined, and detected by a 260nm ultraviolet spectrophotometer, and the absorption peak (the second peak) containing the 146S antigen of the complete viri...

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PUM

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Abstract

The invention relates to an inhibition ELISA (Enzyme-Linked Immunosorbent Assay) method for detecting a foot-and-mouth disease virus antibody and application, and belongs to the technical field of biotechnology. The inhibition ELISA method comprises the following steps of: directly diluting negative and positive serum and detected serum on an ELISA plate coated with a captured antibody, and then adding an equal amount of virus antigen diluted to a working concentration, at the same time, setting at least four-hole virus antigen control (serum diluent with the same volume is added in advance), inhibiting virus antigens from being combined with captured antibodies on an ELISA plate by foot-and-mouth disease type specific antibodies in the detected serum and negative and positive serum, referring to a liquid phase blocking ELISA method for judgment, taking 50% of the average value of OD values of virus antigen control holes as a critical value of 50% inhibition of the virus antigens, thus successfully establishing the inhibition ELISA method for detecting the foot-and-mouth disease antibody.

Description

technical field [0001] The invention belongs to the technical field of biotechnology, relates to a new method for detecting foot-and-mouth disease virus antibody, in particular to an inhibition ELISA method and application for detecting foot-and-mouth disease virus antibody. Background technique [0002] Foot and Mouth Disease (FMD) is an acute, febrile, highly contagious infectious disease that is susceptible to cloven-hoofed animals such as pigs, cattle, and sheep caused by Foot and Mouth Disease Virus (FMDV). Infected animals Most of the clinical features are vesicular eruptions on the oral mucosa, hooves and breast skin. The disease spreads through many routes and is fast. Every outbreak will cause huge economic losses to the local animal husbandry industry. The World Organization for Animal Health (OIE) lists it as an infectious disease that must be declared, and my country also lists it as the first class of animal infectious diseases. Foot-and-mouth disease virus mai...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68G01N33/569G01N33/577G01N33/543
CPCG01N33/6854G01N33/56983G01N33/577G01N33/54393G01N2333/09G01N2469/20
Inventor 周广青马军武邵军军常惠芸龚真莉郭建宏祁淑芸李苗苗何继军
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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